As production methods of L-amino acid, various methods such as extraction procedure, chemical synthetic method, fermentation method, enzymatical synthetic method and the like are known. The extraction procedure requires a large-scale facility for purification of protein hydrolysates. Since amino acid produced by a chemical synthetic method is generally in the form of a racemate, production of an optically active form requires an expensive resolving agent, an asymmetric catalyst and the like. The fermentation method requires a large-scale purification facility as in the extraction procedure since the resulting product has a low concentration, and is unsuitable for the synthesis of non-natural amino acids. The enzymatical synthetic method avoids these problems by utilizing an economical biological catalyst, and can provide an efficient production method at a lower cost.
As an enzymatical synthetic method of L-amino acid, for example, a method including reacting racemic amino acid with D-amino acid oxidase is known.
D-amino acid oxidase (enzyme No. [EC1.4.3.3]) is an enzyme that oxidizes D-amino acid in the presence of oxygen and produces hydrogen peroxide, 2-oxo acid, and ammonia. D-amino acid oxidase is known to have an industrially useful activity such as production of L-amino acid and 2-oxo acid by stereoselective oxidation of racemic amino acid, and the like. In addition, L-amino acid is a useful compound as a synthetic intermediate for pharmaceutical products and the like, and a sweetener.
As microorganisms producing D-amino acid oxidase, for example, Trigonopsis variabilis (Patent Reference 1), Fusarium oxysporum (Patent Reference 2), Candida tropicalis (Non-Patent Reference 1) and the like are known, each of which enzymes was purified and isolated, and the properties thereof have been clarified. In addition, as to the expression of D-amino acid oxidase gene in a transformant, that of genes derived from, for example, Trigonopsis variabilis (Patent Reference 3), Fusarium solani (Patent Reference 4), Rhodotorula gracilis (Patent Reference 5), Candida boidinii (Non-patent Reference 2) are known. For high production of D-amino acid oxidase in these transformants, in many cases, a method of highly producing an enzyme, which includes culturing while suppressing expression, and then adding an inducer and the like, is known.
As regards Candida intermedia belonging to Candida genus, it is known to have a D-amino acid oxidase activity when it is in crude enzyme solution (Non-Patent Reference 3).
Moreover, as an enzymatic synthetic method of L-amino acid, for example, a production method by a stereo inversion reaction, wherein L-amino acid is produced by reacting racemic amino acid with D-amino acid oxidase, amino acid dehydrogenase and an enzyme having a coenzyme-regenerating activity, is known (Non-Patent References 4, 5). However, the methods disclosed in Non-Patent References 4, 5 have problems for industrial application because it requires an expensive commercially available enzyme and a purified enzyme from a microorganism.
To solve this problem, for example, a method comprising preparing and culturing a transformant having respective genes encoding D-amino acid oxidase, amino acid dehydrogenase, an enzyme having coenzyme-regenerating activity, and reacting same with racemic amino acid to give L-amino acid is known (Patent Reference 6). According to the Examples described in Patent Reference 6, however, the concentration of L-amino acid produced by a reaction using the transformant is about 25 mM, which is low for an industrial production.    Patent Reference 1: JP-B-62-501677    Patent Reference 2: JP-A-11-318439    Patent Reference 3: JP-A-63-71180    Patent Reference 4: EP-A-364275    Patent Reference 5: WO97/040171    Patent Reference 6: WO05/090950    Non-Patent Reference 1: “Some Properties of Peroxisome associated D-amino Acid Oxidase from Candida tropicalis”, Agricul. and Biol. Chemistry, 1986, Vol. 50, No. 10, page 2637    Non-Patent Reference 2: “Characterization and High-level Production of D-Amino Acid Oxidase in Candida boidinii”, Biosci. Biotechnol. Biochem., 2001, Vol. 65, No. 3, page 627    Non-Patent Reference 3: “Production of D-Amino Acid Oxidase by Candida tropicalis”, J. Ferment. Technol., 1977, Vol. 55, No. 1, page 13    Non-Patent Reference 4: “Enzymatic Conversion of Racemic Methionine to the L-Enantiomer”, J. Chem. Soc. Chem. Commun., 1990, Vol. 13, page 947    Non-Patent Reference 5: “Enzymatic synthesis of chiral intermediates for Omapatrilat, an antihypertensive drug”, Biomolecular Engineering, 2001, Vol. 17, page 167